The long-term objective of this proposal is to identify the steps linking fuel metabolism and initiation of insulin secretion. In particular, the focus will be the mechanisms involved in regulation of the beta-cell energy state by fuel molecules since insulin secretion is an energy-dependent process and new data suggest that intracellular free Ca2+, the final signal for insulin secretion, is regulated, in part, by the interplay of bioenergetic factors. The specific aims of this project are: (1) to determine the relationship between the beta-cell energy state and insulin secretion by establishing effects of fuel molecules on the energy state and the effect of pharmacologically altering the energy state on insulin secretion. The role of guanine nucleotides in insulin secretion will be determined using similar techniques since they may alter Ca2+ metabolism. (2) to establish the biochemical mechanisms involved in regulation of the energy state by fuel molecules, especially glucose. In particular, we will focus on the role of the alpha-glycerol phosphate shuttle and Ca2+-dependent mitochondrial dehydrogenases in regulating the intramitochondrial redox state. (3) to determine whether physiologic compartmentation of ATP occurs in the cytoplasm of beta-cells and thus whether effector mechanism involved in initiating insulin secretion may respond to localized concentrations of adenine nucleotides. This plan is directed toward a long-term goal of understanding the impairment in beta-cell function occurring in non-insulin dependent diabetes in humans. The elucidation of the regulatory mechanisms involved in fuel metabolism in beta-cells is the basis for understanding the relationship between fuel sensing by beta- cells and stimulus-secretion coupling initiating insulin secretion.